This project was initiated to study the molecular genetics of cell cycle associated genes in C. neoformans. In 2006-2007, we deleted the TUP1 gene, a fungal global repressor known to regulate the rate of cell proliferation, mating and morphogenesis, in H99, a serotype A strain of C. neoformans. The H99 strain is of the VN1 molecular type which significantly differs from serotype D strains in capsular antigenicity and the function of various genes. Unlike in serotype D strains, deletion of the tup1 gene in H99 did not cause a quorum sensning phenotype but the growth on complex media was clearly retarded compared to the wild type strains. Tup1 ko isolates of H99 showed a drastic increase in capsule formation. Complementation of the tup1 ko strain with the wild type TUP1 gene decreased the capsule size significantly. These findings indicate that TUP1 represses capsule formation in vitro. The capsule size in vivo, however, was not affected by the TUP1 deletion. Mice infected with the tup1 ko strain survived much longer than either the wild type or the tup1 ko strain complemented with the TUP1 gene. In the same period, we carried out microarray experiments to compare the differences in the expression of known or novel capsule related genes between the wild type and the tup1 ko strain.Interestingly,the microarray data indicated that expression of multiple genes related to iron homeostasis were affected by the deletion of the tup1 gene. This could explain the cause of the observed hypercapsular phenotype of the tup1 ko strain in serotype A background. A direct relationship between iron homeostasis and the degree of capsule formation is known to exist in Cryptococcus neoformans. Since tup1 deletion in H99 caused no quorum sensing phenotype, we attempted to determine whether such a phenotype is widespread in strains of serotype A of different molecular types. The reference strains of molecular type VNI, VNII and VNBt were chosen to study in 2007-2008. Contrary to what has been known in strains of serotype D, expression of the TUP1 gene in strains of serotype A, irespective of molecular type, is unrelated to production of the quorum sensing peptides. These findings support the observed genetic diversity between strains of serotype D and A of C. neoformans even though they produce identical disease in humans. During the period of 2008-2009, we found other pleiotropic phenotypes in serotype A tup1 null strains that had not been observed in serotype D strains. These include increased susceptibility to azole drugs, the most widely used antimycotics to treat cryptococcosis, sensitivity to SDS and a reduction in the formation of melanin, one of the major virulence factors in C. neoformans. These results suggested the pathobiological importance of TUP1 in C.neoformans.